Re-evaluating microglia expression profiles using RiboTag and cell isolation strategies
| Autor: | Binyamin Zuckerman, Sigalit Boura-Halfon, Zhana Haimon, Anat Shemer, Alon Volaski, Steffen Jung, Igor Ulitsky, Louise Chappell-Maor, Martin Gericke, Ingo Bechmann, Eyal David, Diana Varol, Johannes Orthgiess, Simon Yona |
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| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Genetically modified mouse Immunology Cell Cre recombinase Mice Transgenic Computational biology Biology Ribosome Article Transcriptome Mice 03 medical and health sciences medicine Animals Immunoprecipitation Immunology and Allergy RNA Messenger Cell isolation Gene Microglia Sequence Analysis RNA Mice Inbred C57BL 030104 developmental biology medicine.anatomical_structure Ribosomes |
| Zdroj: | Nature immunology |
| ISSN: | 1529-2916 1529-2908 |
| DOI: | 10.1038/s41590-018-0110-6 |
| Popis: | Transcriptome profiling is widely used to infer functional states of specific cell types, as well as their responses to stimuli, to define contributions to physiology and pathophysiology. Focusing on microglia, the brain’s macrophages, we report here a side-by-side comparison of classical cell-sorting-based transcriptome sequencing and the ‘RiboTag’ method, which avoids cell retrieval from tissue context and yields translatome sequencing information. Conventional whole-cell microglial transcriptomes were found to be significantly tainted by artifacts introduced by tissue dissociation, cargo contamination and transcripts sequestered from ribosomes. Conversely, our data highlight the added value of RiboTag profiling for assessing the lineage accuracy of Cre recombinase expression in transgenic mice. Collectively, this study indicates method-based biases, reveals observer effects and establishes RiboTag-based translatome profiling as a valuable complement to standard sorting-based profiling strategies. Isolating tissue-resident cells runs the risk of altering their expression profile. Jung and colleagues use a RiboTagging approach to describe the microglial translatome and demonstrate that macrophage extraction from their tissue environment results in significant transcriptional alterations. |
| Databáze: | OpenAIRE |
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